Equine herpesviruses comprise a group of antigenic distinct biological agents which cause a variety of infections in horses, ranging from subclinical to fatal disease.
The Equine herpesvirus is inter alia one of the most common causes of infectious abortions and may account for 15% of all mare abortions that occur during the last six months of pregnancy. Equine herpesvirus type 1 (EHV-1) can cause abortion, perinatal foal mortality, respiratory inflammation and neurologic disease.
Although very similar to EHV-4 (previously classified as EHV-1 subtype 2), EHV-1's main clinical sign is abortion while EHV-4 results in respiratory disease. But both cross over and can be clinically indistinguishable as a disease.
The virus is contracted through inhalation. The respiratory signs can range from severe to inapparent. Abortion occurs primarily in mares over 7 months pregnant, follows infection in 14 to 120 days and can occur suddenly without signs in the mare. The virus infects fetal lung and liver tissue and the mare placental endothelial tissue. Abortion may occur because of direct effects on the fetus or because of placental separation. Near term fetuses may be born alive but due quickly due to lung pathology.
Primary infection of upper respiratory tract of young horses results in a febrile illness which lasts for 8 to 10 days. Immunologically experienced mares may be reinfected via the respiratory tract without disease becoming apparent so that abortion usually occurs without warning. The neurological syndrome is associated with respiratory disease or abortion and can affect animals of either sex at any age, leading to incoordination, weakness and posterior paralyses.
Other EHV viruses are EHV-2 or Equine Cytomegalovirus, which is a ubiquitous antigenically heterogeneous, usually slowly growing group of viruses, causing no-known disease, and EHV-3, the Equine Coital exanthema virus which is the causative agent of a relatively mild pregenital exanthema of both mare and stallion. EHV-1 subtype 2 is now called EHV-4, and is primarily associated with respiratory disease, although sporadic EHV-4 induced abortions have been reported.
The genomic structure of the EHV is similar to that of other alpha herpesviruses, comprising a double stranded linear molecule consisting of two covalently linked segments (UL and US), wherein the US segment is flanked by inverted repeats. Of the EHV, EHV-1 is the most extensively studied. Telford, E.A.R. et al., Virology 189, 304–316 (1992) have published the complete DNA sequence of EHV-1. The genome consists of about 150,000 bp and about 76 distinct genes have been recognised up to now.
Equine herpesviruses in general and EHV-1 in particular, are ubiquitous pathogens in horses. EHV-1 can even cause epidemics of abortion, respiratory tract disease and central nervous system disorders. Prevention of infection with the virus is therefore of major economic importance, because EHV can be a severe threat, especially to horses living in close groups, such as studs.
Current vaccines against these viruses comprise chemically inactivated viruses or attenuated live viruses. However, these require multiple administration and have only a limited efficacy.
Inactivated vaccines generally induce only a low level of immunity. Attenuated live vaccines are thus preferred because they evoke a more long-lasting immune response and are easier to produce. Attenuation can be obtained by serial passages of virulent strains in tissue culture of other hosts than the natural one. However, the strains thus obtained are not very well defined and were considered not to be effected. Furthermore, there is always a risk that the viruses revert to virulence resulting in disease in inoculated animals. For this reason, genetic attenuation was adopted as a novel approach to obtain safe vaccine strains.
Genetic attenuation consists for example of deletion of one or more non-essential genes. Examples are viruses with deletions in the thymidine kinase gene or the gene encoding glycoprotein gE. Both have been used successfully for genetic attenuation of herpesviruses in general, but similar mutations in EHV-1 strongly abolished replication in the host and made the viruses no longer useful for vaccine purposes. The mutations furthermore often did not result in the desired level of attenuation.
It is therefore a first objective of the present invention to provide new EHV virus strains that do not have the drawbacks mentioned above when used in vaccines.
It is a second objective of the invention to provide a method for the more general genetic attenuation of herpesviruses.
The third objective of the invention is to provide for live vaccines that comprise said novel EHV mutant viruses.